Recombinant a-L-iduronidase is only one of many enzymes required for the sequential breakdown of two GAGs, heparan sulfate and dermatan sulfate. After uptake into Hurler fibroblasts, the enzyme works very efficiently to degrade GAG stored in the lysosomes. Complete clearance and normalization of stored GAG can occur with only a few percent of normal a-L-iduronidase levels. If the GAGs are labeled using 35SO4, and the cell extracts treated with boiling 80% ethanol, only GAGs remain insoluble and can be precipitated and purified . Using 35SO4 labeling and the extraction procedure, Hurler cell GAG accumulation can be studied in vitro. When Hurler cells are labeled with 35SO4 and incubated with enzyme at different concentrations, the half-maximal correction of abnormal GAG accumulation occurs at a very low enzyme level of about 1 pM, or 1/1000th of the uptake constant of 1 nM. This result indicates the high potency and efficiency of the uptake process, and the ability of exogenous a-L-iduronidase to act in concert with other endogenous enzymes to cleave accumulated GAG back to normal levels.
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